CAREER: Determining and Tuning the Impact of Low-Level Metal-Based Nanoparticles on Actin Cytoskeleton in Normal and Cancer Cells

纳米材料可能会与人体细胞相互作用，这是由于纳米材料无意中释放到环境中（通过空气和水污染），或者有意将纳米材料用于治疗癌症等疾病。细胞暴露于纳米材料可能产生剂量依赖性的有益或有害影响。这些影响传统上用生物化学测定法如细胞活力和膜完整性测定法来评价。然而，这些方法忽略了纳米材料对细胞肌动蛋白细胞骨架（细胞的“骨结构”）的影响。这个CAREER项目将产生关于低水平金属基纳米颗粒如何影响正常和癌细胞中肌动蛋白细胞骨架的新知识。该项目的潜在影响将是为纳米材料安全性评估提供一个基于细胞肌动蛋白细胞骨架的敏感和定量平台，从而能够建立安全和可持续纳米材料的监管和制造框架。该项目还可能导致使用纳米材料治疗癌症转移（90%的人类癌症死亡原因）的安全，有效和通用的方法。该项目的研究综合教育计划非常重视教育第一代大学生和美洲土著学生，这是南达科他州目前辍学率很高的两个代表性不足的群体。通过精心策划的暑期研究经验，预计这两组共10名学生将获得化学，生物学，材料科学和纳米生物技术领域的宝贵实践技能，并增强完成STEM教育的学术坚持性。PI还将在部落学院举办一系列题为“SEE NANO”的研讨会，并将视觉上吸引人和科学上有趣的研究成果与翻转课堂方法相结合，使STEM课堂更加愉快和吸引人，从而促进STEM领域的积极学习和保留。大多数关于纳米材料细胞毒性的研究依赖于宏观生物化学测定，包括细胞活力和膜完整性测定，而没有考虑到与纳米材料的纳米级相互作用，即使在低水平，也可以改变细胞的肌动蛋白细胞骨架。当细胞活力和膜完整性没有改变时，关于细胞的肌动蛋白细胞骨架如何受到低浓度纳米材料的影响存在知识缺口。该CAREER项目的总体研究目标是确定低水平金属基纳米颗粒如何影响正常和癌细胞中的肌动蛋白细胞骨架，并应用这些知识来调整它们对肌动蛋白细胞骨架的影响，以抑制癌细胞迁移而不影响正常细胞。首要的假设是，基于金属的纳米颗粒与肌动蛋白细胞骨架的相互作用机制将取决于它们的表面功能和细胞内降解性，并且这些特性可以被调节以选择性地破坏癌细胞中的肌动蛋白细胞骨架而不影响正常细胞中的肌动蛋白细胞骨架。具体而言，本项目将重点关注两种模型金属基纳米颗粒，包括金属有机框架和金纳米颗粒。目标1将确定低水平的金属基纳米颗粒如何影响血管细胞的肌动蛋白结构和功能。目的2揭示纳米粒子改变癌细胞肌动蛋白结构和迁移能力的机制。最后，目标3将通过将原子力显微镜技术与纳米颗粒的表面功能化相结合，预测和调整纳米材料对血管和癌细胞中肌动蛋白细胞骨架的影响。纳米颗粒引起的肌动蛋白细胞骨架的变化将通过纳米机械原子力显微镜和超分辨荧光显微镜来解决。从这项研究中产生的机制知识将导致：（i）基于血管细胞的生物物理反应的纳米颗粒安全性评估的敏感平台;和（ii）几种通用，有效和安全的方法来阻止癌症转移。该项目的教育目标是提高本科生在STEM教育中的保留率，方法是让一年级学生参与专注于纳米生物相互作用的特定研究任务，在部落学院举办题为“SEE NANO”的系列研讨会，并将翻转课堂方法引入STEM课堂。研究综合教育计划非常重视教育第一代大学生和美洲土著学生，这两个在南达科他州代表性不足的群体在第一年后表现出高辍学率。让这些学生在第一个夏天从事研究将激励和促进他们在完成STEM教育的学术坚持。PI还将整合视觉吸引力和科学有趣的研究成果与翻转课堂方法，使STEM课堂更加愉快和吸引人，从而促进STEM领域的积极学习和保留。该奖项反映了NSF的法定使命，并通过使用基金会的智力价值和更广泛的影响力审查标准进行评估，被认为值得支持。

Nanomaterials may interact with human cells, due to the unintended release of nanomaterials into the environment (through air and water pollution), or intentionally in applications of nanomaterials for treating diseases such as cancer. Exposure of cells to nanomaterials can have dose-dependent beneficial or harmful effects. These effects are traditionally evaluated with biochemical assays such as cell viability and membrane integrity assays. However, these approaches neglect the impact of nanomaterials on the cell’s actin cytoskeleton, the “bone structure” of the cells. This CAREER project will generate new knowledge about how low-level metal-based nanoparticles impact the actin cytoskeleton in normal and cancer cells. The potential impact of this project will be in offering a sensitive and quantitative platform based on actin cytoskeleton of cells for nanomaterial safety assessment, thus enabling the establishment of regulatory and manufacturing frameworks for safe and sustainable nanomaterials. The project will also potentially lead to safe, effective, and versatile approaches for using nanomaterials to treat cancer metastasis (the cause of 90% of human cancer deaths). The research-integrated educational plan in this project places a strong emphasis on educating First-Generation College Students and Native American Students, two underrepresented groups in South Dakota currently exhibiting high dropout rates. Through well-planned summer research experiences, a total of 10 students from these two groups are expected to acquire valuable hands-on skills in the fields of chemistry, biology, materials science, and nanobiotechnology, as well as enhanced academic persistence in completing their STEM education. The PI will also conduct a workshop series entitled “SEE NANO” at Tribal Colleges and integrate visually appealing and scientifically interesting research results with Flipped Classroom Methodology to make STEM classrooms more enjoyable and engaging, thereby promoting active learning and retention in STEM fields.Most research on nanomaterial cytotoxicity relies on macroscopic biochemical assays, including cell viability and membrane integrity assays, without considering that nanoscale interactions with nanomaterials, even at low levels, can alter the actin cytoskeleton of cells. There is a knowledge gap concerning how the actin cytoskeleton of cells is affected by low concentrations of nanomaterials, when cell viability and membrane integrity have not been changed. The overall research goal of this CAREER project is to determine how low-level metal-based nanoparticles affect the actin cytoskeleton in normal and cancer cells, and to apply this knowledge to tune their impact on the actin cytoskeleton for inhibition of cancer cell migration without impacting normal cells. The overarching hypothesis is that the interaction mechanism of metal-based nanoparticles with actin cytoskeleton will be dependent on their surface functionality and intracellular degradability, and these properties can be tuned to selectively disrupt the actin cytoskeleton in cancer cells without affecting the actin cytoskeleton in normal cells. Specifically, this project will focus on two model metal-based nanoparticles including metal-organic frameworks and gold nanoparticles. Objective 1 will determine how low-level metal-based nanoparticles affect the actin structures and functions of vascular cells. Objective 2 will reveal the mechanisms of how these nanoparticles alter the actin structures and migration ability of cancer cells. Finally, Objective 3 will predict and tune the impact of nanomaterials on the actin cytoskeleton in vascular and cancer cells by combining atomic force microscopy techniques with surface functionalization of nanoparticles. The actin cytoskeleton changes induced by the nanoparticles will be resolved by nanomechanical atomic force microscopy and super resolution fluorescence microscopy. The mechanistic knowledge generated from this research will lead to: (i) a sensitive platform for nanoparticle safety assessment that is based on the biophysical responses of vascular cells; and (ii) several versatile, effective, and safe approaches for stopping cancer metastasis. The educational objective of this project is to improve the retention of undergraduates in STEM education, by engaging first-year students to specific research tasks focusing on nano-bio interactions, conducting a workshop series entitled “SEE NANO” at Tribal Colleges, and bringing Flipped Classroom Methodology to STEM classrooms. The research integrated educational plan places a strong emphasis on educating First-Generation College Students and Native American Students, two underrepresented groups in South Dakota exhibiting high dropout rates after their first year. Engaging these students in research during their first summer will motivate and promote their academic persistence in completing STEM education. The PI will also integrate visually appealing and scientifically interesting research results with Flipped Classroom Methodology to make STEM classrooms more enjoyable and engaging, thereby promoting active learning and retention in STEM fields.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.